Role for ICP0 and UL55/UL13/Us10 in protein degradation in neurons and reactivation of HSV from latent infection - The human pathogen, Herpes Simplex Virus type 1 (HSV-1), establishes a latent infection only in neurons and can reactivate to result in the production of infectious virus for transmission to a new host. Reactivation can result in lesions at the body surface and encephalitis, in additional to potentially promoting the development of neurodegenerative disease. HSV-1 has mechanisms to degrade certain host proteins that restrict virus replication in non-neuronal cells, which permits the virus to overcome intrinsic host immunity mediated by restriction factors. However, it is currently unknown what proteins are degraded by the virus in neurons. Understanding proteins targeted for degradation in neurons is important to identify proteins that restrict the virus for either entry into latency or form a barrier to reactivation. One viral protein, ICP0, is an E3 ubiquitin ligases that mediates degradation of host proteins involved in restricting replication. Surprisingly, we have identified a complex containing pUL55, pUL13, and pUS10 that acts redundantly with ICP0 to degrade host proteins in non-neuronal cells. We hypothesize that ICP0 and pUL55/pUL13/pUS10 target specific host proteins for degradation in neurons following HSV-1 infection to enable viral gene expression and modulate the latency/reactivation cycle. The goals of this project are to identify host proteins degraded in neurons and the contribution of the pUL55 complex and ICP0 to both protein degradation in neurons and reactivation. Based on our preliminary data, we hypothesize that ICP0 and pUL55 function independently in neurons to promote viral gene expression. We will use the expertise of the Cliffe lab to investigate HSV neuronal infection, along with the Crump lab to carry out quantitative tandem mass spectrometry on infected cells. In aim 1, using both murine and human neurons, we will determine proteins degraded following HSV-1 neuronal infection and the role of ICP0 and the pUL55 complex and their potential role in restricting neuronal infection. In aim 2, we will use our validated model of HSV latency to determine the exact contribution of ICP0 and pUL55/pUL13/pUS10 to reactivation, using both deletion viruses in addition to shRNA depletion of the proteins to understand their role solely in reactivation. Therefore, this proposal is significant and innovative because it will create a novel dataset of proteins degraded in neurons and test the hypothesis that HSV has evolved multiple strategies to degrade host proteins in neurons. In addition, we will determine the contribution of both ICP0 and the pUL55 complex solely in neuronal infection and reactivation. Overall, these aims will provide insights into the mechanisms of HSV-1 latency in neurons and may lead to the development of novel therapeutic strategies to prevent reactivation from occurring.
